This research aims to elucidate the biochemical pathways that regulate intracellular folate concentrations through the degradation of folate cofactors, a process known as folate catabolism. Folate is a metabolic cofactor that is required for the synthesis of nucleotides and s-adenosylmethionine. These products are key metabolites for DNA synthesis, DNA methylation, protein synthesis, and cell signaling. Disruption of folate metabolism by vitamin deficiency, genetic predisposition or medical therapies leads to elevations in serum homocysteine and increased uracil content in DNA, which are biomarkers that indicate risk for tissue- and cell-specific pathologies including neural tube defects and cancer. Increased rates of folate catabolism occur during states of rapid cell proliferation (i.e. pregnancy and cancer) and can result in localized cellular folate deficiency in the absence of dietary or whole-body deficiency. Recently, our laboratory purified an enzyme that catalyzes the oxidative cleavage of folate coenzymes to inactive degradation products, a reaction that can regulate intracellular folate concentrations in cell cultures. The protein was identified as heavy chain ferritin (HCF). More recent studies have shown that methenyltetrahydrofolate synthetase (MTHFS) also accelerates folate turnover in cells. In this proposal, the biochemical pathways whereby HCF and MTHFS influence folate turnover will be elucidated, and their role in regulating cellular folate concentrations and folate metabolism will be determined in mouse models. The principle hypotheses to be tested are that: (1). folate catabolism is a regulated, enzyme catalyzed reaction in vivo.(2). folate catabolism plays a primary role in regulating intracellular folate concentrations.(3). folate catabolism induces tissue-specific folate deficiency in the absence of whole-body deficiency.The long-term goals of this project are: (1). to elucidate the role(s) of folate catabolism in the regulation of folate metabolic reactions. (2). to determine the risk that folate catabolism contributes to folate-associated pathologies. (3). to establish relationships between catecholamines and folate metabolism.